Identity of the cholesterol-raising factor from boiled coffee and its effects on liver function enzymes.

Boiled coffee contains an unidentified lipid that raises serum cholesterol. We studied the effects of the ingestion of coffee oil fractions of increasing purity in volunteers in order to identify the cholesterol-raising factor. In 15 volunteers who ingested 0.75 g/d of a non-triglyceride-fraction from coffee oil for 4 weeks, mean cholesterol increased by 48 mg/dl (1.2 mmol/l) relative to placebo. In contrast, a coffee oil stripped of the non-triglyceride lipids cafestol and kahweol had no effect. In three volunteers, purified cafestol (73 mg/d) plus kahweol (58 mg/d) increased cholesterol by 66 mg/dl (1.7 mmol/l) after 6 weeks. Oil from Robusta beans, which contains cafestol but negligible kahweol, also raised serum cholesterol. These findings show that cafestol is at least partly responsible for the cholesterol-raising effect of boiled coffee. Coffee oils and brews containing cafestol consistently increased serum triglycerides and alanine amino-transferase, and depressed serum creatinine and gamma-glutamyl-transferase (GGT). After withdrawal GGT activity rose above baseline. Norwegians who habitually consumed 5-9 cups of boiled coffee per day had higher serum cholesterol levels and lower GGT but no higher alanine aminotransferase activity than controls. Thus, serum cholesterol is raised by cafestol and possibly also kahweol, both natural components of coffee beans. The mechanism of action is unknown but is accompanied by alterations in liver function enzymes.

Contribution of coffee aroma constituents to the mutagenicity of coffee.

About 40 coffee aroma constituents belonging to the classes of dicarbonyls, sulphur-containing compounds, furfuryls, N-heterocyclics and others were systematically evaluated in three Ames tester strains. Only aliphatic dicarbonyl compounds showed notable direct mutagenic activity, which mainly affected 'base-pair substitution' in Ames tester strains TA100 and TA102. Very weak effects were also seen with some N-heterocyclics, mainly affecting frameshift tester strain TA98 upon metabolic activation. However, it was shown that these N-heterocyclics do not contribute substantially to the mutagenicity in coffee. The hydrogen peroxide and methylglyoxal contents of coffee were determined up to 26 hr after preparation. Their concentrations tended to decrease whereas mutagenic activity decreased significantly with time in tester strains TA100 and TA102. It is concluded that several highly labile coffee constituents contribute to the bacterial mutagenicity and also that the synergism between hydrogen peroxide and methylglyoxal is not the main factor. The absence of coffee mutagenicity/carcinogenicity in rodents with these highly reactive coffee aroma compounds can be explained in part by detoxification of microsomal enzyme systems.

Stability of tryptophan during food processing and storage. 1. Comparative losses of tryptophan, lysine and methionine in different model systems.

The stability of tryptophan was evaluated in several different food model systems using a chemical method (high pressure liquid chromatography after alkaline-hydrolysis) and rat assays. Losses of tryptophan were compared with the losses of lysine and methionine. Whey proteins stored in the presence of oxidizing lipids showed large losses of lysine and extensive methionine oxidation but only minor losses of tryptophan as measured chemically. The observed decrease in bioavailable tryptophan was explained by a lower protein digestibility. Casein treated with hydrogen peroxide to oxidize all methionine to methionine sulphoxide showed a 9% loss in bioavailable tryptophan. When casein was reacted with caffeic acid at pH 7 in the presence of monophenol monooxygenase (tyrosinase; EC 1.14.18.1), no chemical loss of tryptophan occurred, although fluorodinitrobenzene-reactive lysine fell by 23%. Tryptophan bioavailability fell 15%, partly due to an 8% reduction in protein digestibility. Alkali-treated casein (0.15 M-sodium hydroxide, 80 degrees, 4 h) did not support rat growth. Chemically-determined tryptophan, available tryptophan and true nitrogen digestibility fell 10, 46 and 23% respectively. Racemization of tryptophan was found to be 10% (D/(D+L)). In whole-milk powder, which had undergone "early' or "advanced' Maillard reactions, tryptophan, determined chemically or in rat assays, was virtually unchanged. Extensive lysine losses occurred. It was concluded that losses of tryptophan during food processing and storage are small and of only minor nutritional importance, especially when compared with much larger losses of lysine and the more extensive oxidation of methionine.

Breakfast meal composition influences plasma tryptophan to large neutral amino acid ratios of healthy lean young men.

The effect of a carbohydrate, a 20% protein, or a carbohydrate +0.3% tryptophan TRP breakfast on plasma large neutral amino acid ratios was studied in 6 healthy men. The carbohydrate-rich meal produced shifts in plasma amino acid concentrations such that plasma TRP/LNAA ratios increased from 0.13 to 0.15 (p less than 0.04) and the protein meal decreased the ratio from 0.14 to 0.11 (p less than 0.04) after 1 hour. Addition of 0.3% TRP to the carbohydrate-rich meal increased plasma TRP/LNAA ratios more than 2-fold. The TRP containing meal was thus the only one likely to influence brain 5-HT synthesis, although the difference between the plasma TRP/LNAA ratios after carbohydrate and protein breakfasts suggests that the brain may distinguish, by synthesizing more or less 5-HT, the composition of breakfast meals.

Adipose tissue labelling in man, using a structurally-labelled fatty acid as tracer.

Three healthy male subjects had their adipose tissue enriched with a structurally-labelled fatty acid, 13-methyltetradecanoic acid (13-MTD), occurring naturally only in trace amounts. Needle biopsies from femoral, gluteal and abdominal adipose tissue were obtained at several intervals during a 6-week 13-MTD ingestion period and also along an 11-week period following the cessation of the label intake. Fatty acid composition was analysed by gas-liquid chromatography. Using this method, it was possible to show that different adipose tissues had various responsiveness to labelling (with abdominal fat having the highest one) and hence had different turnover rates. Incorporation and particularly disappearance rates were slow, half-lifes for label incorporation ranged from 2 to 12 weeks and that for label mobilization ranged from 4 to 10 months, thus confirming few other published results. The unexpected observation that incorporation of the label did not cease at the end of the 13-MTD feeding period, but did continue for at least a fortnight, might indicate that existence of a buffer pool where fatty acids may be momentarily stored before their definite incorporation into adipose tissue, as suggested in few other studies.

Racemization of free and protein-bound amino acids in strong mineral acid.

A new GC/MS technique was applied to measure the racemization of amino acids in strong mineral acid. The inversion rate constants of 15 free amino acids were determined under standard hydrolysis conditions (110 degrees, 6 N HCl in evacuated vacuum-sealed tubes). The variations in the inversion rate constants of all tested amino acids with the exception of serine and threonine were related to two factors: side-chain electron-withdrawing capacity ( sigma ) and steric hindrance in the vicinity of the alpha-hydrogen atom. The rate constants for serine and threonine were much lower than expected. Hydrolysis-induced racemization of protein-bound amino acids was investigated with alpha-lactalbumin and beta-lactoglobulin. Significant differences were observed as compared with the racemization rates of free amino acids. Such discrepancies were also observed between the two proteins. In the early stage of alpha-lactalbumin hydrolysis, 10% inversion of methionine was measured as compared with less than 0.5% in the case of beta-lactoglobulin. We attributed the particular behaviour of methionine in alpha-lactalbumin to the neighborhood of a cysteine residue on the carboxyl side.

Mass spectra of alpha-amino acid oxazolidinones.

2-Bis-(chlorodifluoromethyl)-4-substituted-1,3-oxazolidin-5-ones, a new type of alpha-amino acid derivative for gas chromatographic separation, have been studied by low resolution mass spectrometry. These derivatives are obtained by reacting alpha-amino acids with dichlorotetrafluoroacetone. Their structure has been established or confirmed for most protein amino acids and several non-protein alpha-amino acids. The mechanisms responsible for the mass spectral pattern have been rationalized. An interesting feature of this derivatization procedure is that it distinguishes aspartic and glutamic acid from the respective amides. The structure of asparagine and glutamine derivatives has been established. A survey of oxazolidinone mass spectra has provided a list of diagnostically useful ions. Each amino acid can be identified by one or two of its most abundant fragments.